31:	Identifying genes involved with floral induction in Perilla crispa using nonradioactive differential display.

Authors:	Cooper, Lol, L.D.(A)Doss, Robert, P.(A)
Affiliations:	(A): Horticultural Crops Research Unit, USDA-ARS, Corvallis, Oregon
Presenter:	Cooper, Lol L.D., cooperl@bcc.orst.edu

A nonradioactive version of differential display was used to screen messenger RNA from photoperiodically induced and noninduced leaves of Perilla crispa, a qualitative short day plant. cDNA populations, generated using oligo (dT)12-18 primers, were screened using combinations of anchored primers and arbitrary decamers. The amplification products were electrophoresed using 6% nondenaturing PAGE and visualized by silver staining. One primer combination yielded a differentially displayed band in a series of independent cDNA replicates. The differentially displayed band was isolated and the cDNA was cloned. Transformants were screened by colony PCR and by restriction digestion of plasmid DNA. Ten transformants with correctly-sized inserts were found to constitute four consensus sequence groups. Unique restriction sites in each of the consensus sequences were used to identify the differentially displayed sequence. Sequence-specific primers were designed to amplify the cDNA clone giving rise to the differentially displayed band. To determine whether or not the cloned sequence is induction-specific, mRNA levels will be compared in photoperiodically induced and noninduced leaves using competitive RT-PCR and northern blot analysis.